Plants, like all higher eukaryotes, must control the onset and spread of cell death during development and in response to environmental signals. Plant biology is replete with examples of developmentally programmed cell death (PCD). A specialized form of PCD in plants, termed the Hypersensitive Response (HR) is tightly correlated with successful recognition of, and response to, pathogen infection. The HR is an important part of the plant immune response. HR is associated with an oxidative burst and signaling of pro-and anti-death signals to cells surrounding the infection site. While much is known about the molecular mechanisms of PCD control in animals, very little is known in plants. This proposal uses Arabidopsis as a genetic model with which to understand the control of HR as a paradigm for PCD in plants. The Arabidopsis genome is fully sequenced and there is little molecular evidence for significant conservation of key regulators of animal PCD at the sequence level. We were among the first to recognize the use of Arabidopsis to genetically dissect cell death control and have made significant contributions to the field. We identified and analyzed a series of mutants that mis-regulate HR-like cell death in the absence of pathogen. We cloned three key HR regulators all belonging to one gene family: LSD1 and the related genes LOL1 (LSD One Like 1) and LOL2. LSD1 acts to suppress the spread of unwanted cell death following a normal HR. Superoxide derived from a plasma membrane NADPH oxidase acts in concert with LSD1 in this process. We recently demonstrated that LSD1 interacts with several proteins, including functionally relevant transcription factors (TFs) and putative "metacaspases". We demonstrated that one TF of the bZIP class, functions in HR and in basal defense to infection. Its activity is antagonized by LSD1, and they interact with in vivo. We intend to characterize the role of the other LSD-interacting TFs in cell death and HR in the coming proposal period. We also recently cloned a second suppressor of the idiosyncratic runaway cell death phenotype of the Isd1 mutant. Very surprisingly, this suppressor encodes a disease resistance protein of the NB-LRR class. This is the first time that an NB-LRR class protein has been implicated in any process other than pathogen recognition. We will investigate how this particular NB-LRR controls the spread of HR-like cell death. We also recently demonstrated that two of the three so-called "metacaspases," identified in Arabidopsis by informatics approaches, and carrying the zinc-finger domain that defines the LSD1 protein family, also function in HR and runaway cell death is Isd1. This is the first definition of a function for these proteins, and allows us to propose a detailed characterization of their action as positive cell death regulators.